The twisting and bending of DNA is an extensively studied aspect of its structure. It affects both structural transitions and interactions between DNA and other molecular complexes. For example, a locally underwound DNA is necessary for transcriptional activation and recombinational repair. Supercoiled DNA is also a key structural factor in chromosomal organization in which the winding of the molecule around histone proteins is necessary for DNA compaction. More specifically, the entropic tension generated in supercoiled DNA in anaphase during chromosomal condensation is released by the action of a specific enzyme, topoisomerase II, thus allowing the disentanglement and segregation of the chromosomes necessary before cell division.
In the last decade new tools (atomic-force-microscopy, optical tweezers, small glass fibers, . . . ) have been developed to manipulate small objects and also to investigate the forces involved in the systems studied.
[1] S. B. Smith, Y. Cui, C. Bustamante Science 271, 795 (1996)
[2] P. Cluzel, A. Lebrun, C. Heller, R. Lavery, J. -L. Viovy, D. Chatenay, F. Caron Science 271, 795 (1996).
Spectacular results were obtained on molecules such as DNA and various motor proteins : RNA polymerase, F1-ATP synthase and myosin, for example.
However, sophisticated instrumentation is required in most of these systems.
The glass fibers have the advantage of giving very quick results. However, they require one to calibrate their elasticity before any measurements are made and although they are able to measure forces stronger than Brownian ones, they are not sensitive enough to be used in the entropic regime (<1 pN).
The AFM may be used in the same way and has the same drawbacks.
Optical tweezers have also been used, for example to measure the force (—˜ 6 pN) produced by a single myosin on an actin filament (the two basic components of muscles). They also require a force calibration. One needs to know the relation between the intensity of the laser beam and the force applied to the system, and one has to determine it every time one changes the trapped object.
Further drawbacks of optical tweezers are the lack of total torsionnal control on DNA and the local heating of the solution by the focussed laser which increases the noise.
Another technique which has been proposed in
[3] Smith S. B., Finzi L., Bustamante C.—Science, vol. 258, Nov. 13 1992, consists in chemically attaching the DNA molecules by one end to a glass surface and by the other end to a magnetic bead. The beads were manipulated by using magnetic and hydrodynamic forces and observed under an optical microscope. Extension versus force curves were obtained.
However, the apparatus and methods proposed in this article did not allow for twisting the molecules.
Also, sophisticated means were needed to infer the forces applied to the beads.